CHAPTER UNDER CONSTRUCTION

Summary

The Corticoviridae is a family of icosahedral internal membrane-containing viruses with double-stranded circular DNA genomes of ~10 000 bases in length. Only one virus species, Pseudoalteromonas virus PM2, has been recognized. Bacteriophage PM2 infects Gram-negative bacteria. Currently, PM2 does not share any significant sequence similarity to any other virus. PM2 was isolated from seawater in 1968 and was already reported in the 1st Report of ICTV. PM2 is the first bacteriophage in which the presence of lipids in the virion was demonstrated. Since only one genus Corticovirus is recognized, the family description corresponds to the genus description.

Virion

Morphology

Icosahedral virions consist of an internal membrane and an outer protein capsid that has a diameter of 57 nm between facets (Figure 1.Corticoviridae) [{Abrescia et al., 2008:18775333RJOHTXAbrescia et al., 2008, Insights into virus evolution and membrane biogenesis from the structure of the marine lipid-containing bacteriophage PM2, Mol Cell, 31, 5, 749-61}]. The genome is enclosed by the membrane. The capsid consists of 200 major capsid protein P2 trimers that are organized on a pseudo T=21 lattice [{Abrescia et al., 2008:18775333RJOHTXAbrescia et al., 2008, Insights into virus evolution and membrane biogenesis from the structure of the marine lipid-containing bacteriophage PM2, Mol Cell, 31, 5, 749-61}]. Protein P2 is composed of two beta-barrels disposed normal to the capsid surface. The P2 trimers have pseudo-six-fold symmetry and the structure is stabilized by calcium ions. Spikes protrude about 8 nm from the capsid surface at the five-fold vertices. The spikes are homopentamers and formed of protein P1. P1 is composed of three beta-barrel domains arranged end to end. The distal C-terminal domains of P1 are used for receptor recognition. The N-termini of P1 form pentagonal assemblies at the vertices. The inner membrane (47 nm in diameter) contains host plasma membrane-derived phospholipids and virus-encoded proteins P3 to P10 [{Kivelä et al., 2002:12134022RJOHTXKivelä et al., 2002, Bacteriophage PM2 has a protein capsid surrounding a spherical proteinaceous lipid core, J Virol, 76, 16, 8169-78}]. Transmembrane proteins P3 and P6 are organized into a lattice on the membrane vesicle surface, on which the outer protein capsid is assembled [{Abrescia et al., 2008:18775333RJOHTXAbrescia et al., 2008, Insights into virus evolution and membrane biogenesis from the structure of the marine lipid-containing bacteriophage PM2, Mol Cell, 31, 5, 749-61}].

Nucleic acid

The genome is a highly supercoiled circular dsDNA of 10,079 bp (6.6×106 Da). DNA comprises about 14% of the virion weight and the G+C content is 42.2%. The phage PM2 genome has been sequenced [{Männistö et al., 1999:10502514RJOHTXMännistö et al., 1999, The complete genome sequence of PM2, the first lipid-containing bacterial virus to be isolated, Virology, 262, 2, 355-63}](AF155037).

Proteins

The genome has 21 putative genes, 17 of which have been shown to code for structural proteins (P1–P10) and nonstructural proteins (P12–P18; Table 2.Corticoviridae). Proteins form about 72% of the virus particle weight.

Table 2.Corticoviridae. Pseudoalteromonas phage PM2 proteins

Proteina

Mass (kDa)

Location/functionb

P1

37.5

Spike protein, receptor binding (S)

P2

30.2

Major capsid protein (S)

P3

10.8

Major membrane protein (S)

P4

4.4

Membrane (S)

P5

17.9

Membrane (S)

P6

14.3

Major membrane protein (S)

P7

3.6

Membrane (S)

P8

7.3

Membrane (S)

P9

24.7

Putative packaging ATPase (S)

P10

29.0

Membrane (S)

P12

73.4

Replication initiation protein (N)

P13

7.2

Transcription factor (N)

P14

11.0

Transcription factor (N)

P15

18.1

Regulative function (N)

P16

10.3

Regulative function (N)

P17

6.0

Lysis factor (N)

P18

5.7

Lysis factor (N)

a P is for protein; Arabic numeral corresponds to the Roman numeral of the gene.

b S is for structural protein; N is for non-structural protein.

Lipids

Particles are about 14% lipid by weight [{Camerini-Otero and Franklin 1975:1140192RJOHTXCamerini-Otero and Franklin 1975, Structure and synthesis of a lipid-containing bacteriophage. The molecular weight and other physical properties of bacterophage PM2, Eur J Biochem, 53, 2, 343-8}]. The membrane contains about 34% phosphatidyl ethanolamine and about 66% phosphatidyl glycerol and trace amounts of phosphatidic acid and acyl-phosphatidyl glycerol [{Braunstein and Franklin 1971:4330370RJOHTXBraunstein and Franklin 1971, Structure and synthesis of a lipid-containing bacteriophage. V. Phospholipids of the host BAL-31 and of the bacteriophage PM2, Virology, 43, 3, 685-95RJOMREFCamerini-Otero and Franklin 1972:4559686RJOHTXCamerini-Otero and Franklin 1972, Structure and synthesis of a lipid-containing bacteriophage. XII. The fatty acids and lipid content of bacteriophage PM2, Virology, 49, 2, 385-93RJOMREFTsukagoshi et al., 1976:990298RJOHTXTsukagoshi et al., 1976, Identification of acyl phosphatidylglycerol as a minor phospholipid of Pseudomonas BAL-31, Biochim Biophys Acta, 450, 2, 131-6}]. The lipids are derived from the host plasma membrane, but their composition deviates from that of the host bacterium. Lipids form an internal membrane with virus-specific membrane associated proteins.

Genome organization and replication

To infect and replicate, PM2 delivers its genome across the cell envelope of two known marine host strains: Gram-negative Pseudoalteromonas species ER72M2 and BAL-31. Virions adsorb via the distal tips of the spike proteins to uncharacterized receptors [{Abrescia et al., 2008:18775333RJOHTXAbrescia et al., 2008, Insights into virus evolution and membrane biogenesis from the structure of the marine lipid-containing bacteriophage PM2, Mol Cell, 31, 5, 749-61}]. The internal membrane mediates the translocation of the supercoiled genome across the host cell envelopes most probably via fusion in a process which is not fully understood. Replication of the PM2 genome, most probably by a rolling circle mechanism, takes place in proximity to the host cytoplasmic membrane. The largest PM2 gene XII encoding protein P12 shares significant sequence similarity with the superfamily I group of replication initiation proteins [{Männistö et al., 1999:10502514RJOHTXMännistö et al., 1999, The complete genome sequence of PM2, the first lipid-containing bacterial virus to be isolated, Virology, 262, 2, 355-63}]. The genome is organized in three operons (Figure 2.Corticoviridae). Operons OEL and OER encode early function gene products: the replication initiation protein P12 and transcription regulatory proteins P13, P14, P15 and P16. Expression of the genes for structural proteins is under the control of the late promoter (OL), which is activated by the phage-encoded transcription factors P13 and P14 [{Mannistö et al., 2003:12754225RJOHTXMannistö et al., 2003, Transcription of bacteriophage PM2 involves phage-encoded regulators of heterologous origin, J Bacteriol, 185, 11, 3278-87}]. The mature virions are released from the cell by lysis. Lysis factor P17 permeabilizes the cytoplasmic membrane and acts like a holin, whereas lysis factor P18 disrupt the outer membrane, while peptidoglycan is most probably disrupted by the host lytic factors [{Krupovic et al., 2007:17555443RJOHTXKrupovic et al., 2007, A novel lysis system in PM2, a lipid-containing marine double-stranded DNA bacteriophage, Mol Microbiol, 64, 6, 1635-48}].

Figure 2.Corticoviridae. Genome organization of Pseudoalteromonas phage PM2 (PM2). The genome is a 10,079 bp highly supercoiled circular dsDNA molecule containing 21 ORFs (as arrows showing their orientations) and three operons (OER, OEL and OL). ORFs shown to code for functional proteins are classified as genes and given a Roman numeral. The different colors indicate the ORFs (yellow), a gene for replication initiation protein (orange) and the following groups of genes: transcriptional regulation (magenta), structural proteins (blue) and lysis (green).

Biology

Phages are virulent and replicate in two known strains of marine host bacteria of the genus Pseudoalteromonas. Although PM2 is virulent and the sole member of the family Corticoviridae, using a comparative genomic approach, integrated corticoviral genetic elements have been identified to commonly reside within aquatic bacterial chromosomes [{Krupovic and Bamford 2007:17634101RJOHTXKrupovic and Bamford 2007, Putative prophages related to lytic tailless marine dsDNA phage PM2 are widespread in the genomes of aquatic bacteria, BMC Genomics, 8, 236}].